Figure S1 Second-hand tobacco smoke (SHS) exposure effects on endogenous substance P contribution to spiking response. A. Group data show that spiking response was significantly attenuated by NK1 receptor antagonism in both filtered air (FA) and SHS exposure groups [two-way repeated measures (RM) anova interaction, F(6,66) = 12.606, P < 0.001 and F(6,72) = 4.447, P < 0.001 respectively]. B. NK1 receptor sensitive spiking response to depolarizing current injection. The antagonist effects on spiking responses were not significantly different between FA and SHS exposure groups (two-way RM anova interaction, F(6,138) = 1.612, P = 0.148), masking SHS exposure effect as observed especially in delayed-onset spiking neurons.

Figure S2 Intense substance P immunoreactivity in primate nucleus tractus solitarius (see Appendix S1 for method). Photographs show examples of substance P immunoreactivity in the nucleus tractus solitarius from each one rhesus monkey and one guinea pig. These photographs were obtained under the same settings of microscope, laser and fluorescent detection, as well as the same number of slices used for Z-stack reconstruction between two animal species. Green colour indicates substance P immunoreactivity and blue indicates neuronal specific nuclear protein immunoreactivity. Substance P immunoreactivity is more intense in the rhesus monkey (left) than the guinea pig (middle), confirmed by group data (right; unpaired t-test, *P = 0.022). The number in the parentheses indicates the number of animals.

Figure S3 NK1 receptor antagonist depressed tractus solitarius-evoked excitatory postsynaptic current (EPSC) amplitudes in nucleus tractus solitarius neurons. A. Example traces of tractus solitarius-evoked EPSCs (eEPSCs) from a filtered air (FA)-exposed animal in the absence (gray) or presence (black) of NK1 receptor antagonist (SR140333, 3 µM) under the voltage clamp condition at −60 mV. Dots (•) indicate stimulus artefacts with 20 ms intervals. B. Group data for the first eEPSC amplitude in the absence (solid) and presence (hatched) of NK1 receptor antagonist. NK1 receptor antagonist significantly decreased first eEPSC amplitudes in both FA and Second-hand tobacco smoke (SHS) exposure groups (paired t-test, *P = 0.006 and *P = 0.037 respectively). The effect of the antagonist was not different between exposure groups (unpaired t-test, P = 0.222) (right graph). C. Group data of paired pulse ratio (PPR) in the absence (solid) and presence (hatched) of NK1 receptor antagonist. PPR was calculated by dividing first eEPSC peak amplitudes into second eEPSC peak amplitudes. As the second eEPSC amplitudes were also decreased by NK1 receptor antagonist (paired t-test, P = 0.019 for FA and P = 0.013 for SHS), the paired pulse ratio was not changed by the antagonist in either exposure group (paired t-test, FA: P = 0.135; SHS: P = 0.375) (left graph). The antagonist effects between exposure groups were not different (unpaired t-test, P = 0.151) (right graph).

Appendix S1 Supplemental method.

BPH_1271_sm_Fig_S1-S3_App_S1.doc46KSupporting info item
BPH_1271_sm_Fig_1_LZW.tif824KSupporting info item
BPH_1271_sm_Fig_2_LZW.tif3351KSupporting info item
BPH_1271_sm_Fig3.tif1557KSupporting info item
BPH_1271_sm_Review_Spiking_SP.tif979KSupporting info item
BPH_1271_sm_Review_SYN.tif6562KSupporting info item

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